Drosophila cryopreservation and rewarming for long-term storage ABSTRACT Drosophila melanogaster is one of the leading animal models for biomedical research and development. The importance of Drosophila as a research tool relies on several characteristics: biomedical relevance (> 60% of protein-coding genes in Drosophila have human homologs), ease of culture, small genome size, high fecundity, short generation time, and easily recovered mutations. Currently there are >150K stocks held in centers at Indiana, Vienna, and Kyoto, and this number is growing. However, reliable and cost-effective approaches for long-term preservation of Drosophila stocks are lacking. Methods to cryopreserve Drosophila embryos have been published, but they are considered labor-intensive, have proven difficult to successfully reproduce, and therefore have not been translated into wide practice. Using cryopreservation and rewarming expertise developed over the past two decades in the Bischof lab at the University of Minnesota, we have developed a new and simple way to successfully cryopreserve Drosophila embryos. Our preliminary results are very promising: out of the initial embryos used for cryopreservation, 208% embryos hatched after cryopreservation, and 1.51% embryos developed into fertile adults. For this proposal, our main hypothesis is that we can achieve 10% growth to normal adulthood from cryopreserved Drosophila embryos by optimizing pre-cooling, cooling, warming, and post-warming procedures. In addition to optimizing these procedures for a few established Drosophila lines, we will test them on 10 mutant lines to show that they do not affect mutations.